Project Summary/Abstract Rett Syndrome (RTT) is a currently untreatable disorder caused by loss-of-function mutations in the MECP2 gene and is characterized by uncoordinated or repetitive movements, loss of verbal communication and other cognitive deficits, heightened anxiety, and autonomic and respiratory dysregulation. Mouse models of RTT recapitulate many phenotypes of the human disorder and provide a window into potential therapeutic targets. Although much is understood about the behavioral phenotypes of RTT, little is known about circuit deficits underlying particular behaviors; therefore, the purpose of this study is to use the medial prefrontal cortex (mPFC) as a model for understanding the circuit dysfunction underlying cognitive abnormalities in RTT. Recent data from our lab demonstrates that although Mecp2 mutants are capable of learning cue-shock pairings, they have not consolidated this memory when tested 24 hours later. Additionally, we have shown that the mutant mPFC is hypoexcitable, receiving reduced excitatory drive and exhibiting reduced excitatory synaptic morphology and protein expression. Other labs have demonstrated that memory consolidation in cue- dependent fear conditioning is dependent on activity in the mPFC; given this, we hypothesize that reduced excitability in the Mecp2 mutant mPFC underlies or contributes to the consolidation deficit. Our preliminary data using a chemical-genetic approach to express the excitatory DREADD (Designer Receptor Exclusively Activated by Designer Drugs) suggests that increasing activity, specifically in the mPFC, is capable of reversing this memory deficit. Aim 1 will expand on this finding and further determine which subregion of the mPFC underlies consolidation deficits in Mecp2 mutants. Next we will test two mechanisms which have the potential to rescue activity deficits in the mPFC. Specifically, we have found that hypoactivity in the mPFC of Mecp2 mutants is associated with reductions both in Brain Derived Neurotrophic Factor (BDNF) and in the activity of S6K as measured by the phosphorylation of ribosomal protein S6 (rp-S6), two signaling components known to regulate synaptic connectivity and excitatory dendrite morphology. Therefore, Aim 2 will test the hypothesis that chronic activation of the BDNF receptor TrkB is sufficient to rescue both memory consolidation and excitatory morphology deficits in Mecp2 mutants (using Mecp2 mutants which genetically express green fluorescent protein in the subset of excitatory pyramidal cells that express Thy1; Thy1GFPM) by treating mutants chronically with a TrkB agonist. Finally, Aim 3 will test the hypothesis that constitutive activation of S6K is sufficient to mimic (either completely or partially) the rescue of memory consolidation and excitatory connectivity deficits in Mecp2 mutants observed in Aims 1 and 2. This experiment will use the viral expression of a constitutively active mutant S6K protein specifically in the mPFC of Mecp2:Thy1GFPM mutants. By using the mPFC as a model circuit to address abnormal neuronal activity and S6K activation, both of which are observed throughout the Mecp2 mutant brain, this study has the potential to elucidate mechanisms which underlie the many diverse symptom domains of RTT.